RESUMEN
Diabetes initiates inflammation that can impair the retinal vasculature, and lead to diabetic retinopathy; one of the leading causes of blindness. Inflammatory pathways have been examined as potential therapeutic targets for diabetic retinopathy, but there is still a need for early-stage treatments. We hypothesized that the CD40-TNF Receptor Associated Factor 6 (TRAF6) axis plays a pivotal role in the onset of diabetic retinopathy, and that the CD40-TRAF6 axis would be a prime therapeutic target for early-stage non-proliferative diabetic retinopathy. The CD40-TRAF6 complex can initiate NFκB activation, inflammation, and tissue damage. Further, CD40 and TRAF6 are constitutively expressed on Muller glia, and upregulated in the diabetic retina. Yet the role of the CD40-TRAF6 complex in the onset of diabetic retinopathy is still unclear. In the current study, we examined the CD40-TRAF6 axis in diabetic retinopathy using a small molecule inhibitor (SMI-6877002) on streptozotocin-induced diabetic mice. When CD40-TRAF6-dependent inflammation was inhibited, retinal vascular leakage and capillary degeneration was ameliorated in diabetic mice. Collectively, these data suggest that the CD40-TRAF6 axis plays a pivotal role in the onset of diabetic retinopathy, and could be a novel therapeutic target for early diabetic retinopathy.
Asunto(s)
Diabetes Mellitus Experimental , Retinopatía Diabética , Animales , Ratones , Antígenos CD40/metabolismo , Diabetes Mellitus Experimental/metabolismo , Inflamación/complicaciones , Ratones Endogámicos C57BL , Estreptozocina , Factor 6 Asociado a Receptor de TNF/metabolismoRESUMEN
The global number of diabetics continues to rise annually. As diabetes progresses, almost all of Type I and more than half of Type II diabetics develop diabetic retinopathy. Diabetic retinopathy is a microvascular disease of the retina, and is the leading cause of blindness in the working-age population worldwide. With such a significant health impact, new drugs are required to halt the blinding threat posed by this visual disorder. The cause of diabetic retinopathy is multifactorial, and an optimal therapeutic would halt inflammation, cease photoreceptor cell dysfunction, and ablate vascular impairment. XMD8-92 is a small molecule inhibitor that blocks inflammatory activity downstream of ERK5 (extracellular signal-related kinase 5) and BRD4 (bromodomain 4). ERK5 elicits inflammation, is increased in Type II diabetics, and plays a pathologic role in diabetic nephropathy, while BRD4 induces retinal inflammation and plays a role in retinal degeneration. Further, we provide evidence that suggests both pERK5 and BRD4 expression are increased in the retinas of our STZ (streptozotocin)-induced diabetic mice. Taken together, we hypothesized that XMD8-92 would be a good therapeutic candidate for diabetic retinopathy, and tested XMD8-92 in a murine model of diabetic retinopathy. In the current study, we developed an in vivo treatment regimen by administering one 100 µL subcutaneous injection of saline containing 20 µM of XMD8-92 weekly, to STZ-induced diabetic mice. XMD8-92 treatments significantly decreased diabetes-mediated retinal inflammation, VEGF production, and oxidative stress. Further, XMD8-92 halted the degradation of ZO-1 (zonula occludens-1), which is a tight junction protein associated with vascular permeability in the retina. Finally, XMD8-92 treatment ablated diabetes-mediated vascular leakage and capillary degeneration, which are the clinical hallmarks of non-proliferative diabetic retinopathy. Taken together, this study provides strong evidence that XMD8-92 could be a potentially novel therapeutic for diabetic retinopathy.
RESUMEN
Diabetic retinopathy is the leading cause of blindness in the working-age population worldwide. Although the cause of diabetic retinopathy is multifactorial, IL-17A is a prevalent inflammatory cytokine involved in the promotion of diabetes-mediated retinal inflammation and the progression of diabetic retinopathy. The primary source of IL-17A is Th17 cells, which are T helper cells that have been differentiated by dendritic cells in a proinflammatory cytokine environment. Aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that can manipulate dendritic cell maturation, halt the production of IL-6 (a proinflammatory cytokine), and suppress Th17 cell differentiation. In the current study, we examined the efficacy of an AhR agonist, VAF347, as a potential therapeutic for the onset of non-proliferative diabetic retinopathy in streptozotocin (STZ)-induced diabetic C57BL/6 mice. We determined that diabetes-mediated leukostasis, oxidative stress, and inflammation in the retina of STZ-diabetic mice were all significantly lower when treated with the AhR agonist VAF347. Furthermore, when VAF347 was subcutaneously injected into STZ-diabetic mice, retinal capillary degeneration was ameliorated, which is the hallmark of non-proliferative diabetic retinopathy in this diabetes murine model. Collectively, these findings provide evidence that the AhR agonist VAF347 could be a potentially novel therapeutic for non-proliferative diabetic retinopathy.
Asunto(s)
Diabetes Mellitus Experimental/complicaciones , Retinopatía Diabética/tratamiento farmacológico , Inflamación/tratamiento farmacológico , Pirimidinas/farmacología , Receptores de Hidrocarburo de Aril/agonistas , Animales , Diferenciación Celular , Retinopatía Diabética/etiología , Retinopatía Diabética/metabolismo , Retinopatía Diabética/patología , Inflamación/etiología , Inflamación/metabolismo , Inflamación/patología , Interleucina-17/inmunología , Interleucina-17/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Transducción de SeñalRESUMEN
Diabetic retinopathy is a diabetes-mediated retinal microvascular disease that is the leading cause of blindness in the working-age population worldwide. Interleukin (IL)-17A is an inflammatory cytokine that has been previously shown to play a pivotal role in the promotion and progression of diabetic retinopathy. Retinoic acid-related orphan receptor gammaT (RORγt) is a ligand-dependent transcription factor that mediates IL-17A production. However, the role of RORγt in diabetes-mediated retinal inflammation and capillary degeneration, as well as its potential therapeutic attributes for diabetic retinopathy has not yet been determined. In the current study, we examined retinal inflammation and vascular pathology in streptozotocin-induced diabetic mice. We found RORγt expressing cells in the retinal vasculature of diabetic mice. Further, diabetes-mediated retinal inflammation, oxidative stress, and retinal endothelial cell death were all significantly lower in RORγt-/- mice. Finally, when a RORγt small molecule inhibitor (SR1001) was subcutaneously injected into diabetic mice, retinal inflammation and capillary degeneration were ameliorated. These findings establish a pathologic role for RORγt in the onset of diabetic retinopathy and identify a potentially novel therapeutic for this blinding disease.
Asunto(s)
Capilares/efectos de los fármacos , Diabetes Mellitus Experimental/metabolismo , Retinopatía Diabética/metabolismo , Interleucina-17/metabolismo , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/antagonistas & inhibidores , Vasos Retinianos/metabolismo , Sulfonamidas/farmacología , Tiazoles/farmacología , Animales , Capilares/patología , Muerte Celular/genética , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Diabetes Mellitus Experimental/inducido químicamente , Retinopatía Diabética/inducido químicamente , Retinopatía Diabética/tratamiento farmacológico , Agonismo Inverso de Drogas , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Hiperglucemia/sangre , Hiperglucemia/genética , Inflamación/genética , Inflamación/metabolismo , Inflamación/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/genética , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/metabolismo , Estrés Oxidativo/genética , Vasos Retinianos/efectos de los fármacos , Vasos Retinianos/patología , Sulfonamidas/uso terapéutico , Tiazoles/uso terapéuticoRESUMEN
AIMS/HYPOTHESIS: Levels of neutrophil elastase, a serine protease secreted by neutrophils, are elevated in diabetes. The purpose of this study was to determine whether neutrophil elastase (NE) contributes to the diabetes-induced increase in retinal vascular permeability in mice with streptozotocin-induced diabetes, and, if so, to investigate the potential role of IL-17 in this process. METHODS: In vivo, diabetes was induced in neutrophil elastase-deficient (Elane-/-), Il-17a-/- and wild-type mice. After 8 months of diabetes, Elane-/- mice and wild-type age-matched control mice were injected with FITC-BSA. Fluorescence microscopy was used to assess leakage of FITC-BSA from the retinal vasculature into the neural retina. The level of NE in Il-17a-/- diabetic retina and sera were determined by ELISA. In vitro, the effect of NE on the permeability and viability of human retinal endothelial cells and the expression of junction proteins and adhesion molecules were studied. RESULTS: Eight months of diabetes resulted in increased retinal vascular permeability and levels of NE in retina and plasma of wild-type animals. All of these abnormalities were significantly inhibited in mice lacking the elastase. The diabetes-induced increase in NE was inhibited in mice lacking IL-17. In vitro, NE increased retinal endothelial cell permeability, which was partially inhibited by a myeloid differentiation primary response 88 (MyD88) inhibitor, NF-κB inhibitor, and protease-activated receptor (PAR)2 inhibitor. NE degraded vascular endothelial-cadherin (VE-cadherin) in a concentration-dependent manner. CONCLUSIONS/INTERPRETATION: IL-17 regulates NE expression in diabetes. NE contributes to vascular leakage in diabetic retinopathy, partially through activation of MyD88, NF-κB and PAR2 and degradation of VE-cadherin.
Asunto(s)
Barrera Hematorretinal/metabolismo , Retinopatía Diabética/metabolismo , Elastasa de Leucocito/metabolismo , Retina/metabolismo , Vasos Retinianos/metabolismo , Animales , Barrera Hematorretinal/patología , Permeabilidad Capilar/genética , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Retinopatía Diabética/genética , Retinopatía Diabética/patología , Interleucina-17/genética , Interleucina-17/metabolismo , Elastasa de Leucocito/genética , Masculino , Ratones , Ratones Noqueados , Retina/patología , Vasos Retinianos/patologíaRESUMEN
PURPOSE: Diabetes leads to progressive complications such as diabetic retinopathy, which is the leading cause of blindness within the working-age population worldwide. Interleukin (IL)-17A is a cytokine that promotes and progresses diabetes. The objective of this study was to determine the role of IL-17A in retinal capillary degeneration, and to identify the mechanism that induces retinal endothelial cell death. These are clinically meaningful abnormalities that characterize early-stage non-proliferative diabetic retinopathy. METHODS: Retinal capillary degeneration was examined in vivo using the streptozotocin (STZ) diabetes murine model. Diabetic-hyperglycemia was sustained for an 8-month period in wild type (C57BL/6) and IL-17A-/- mice to elucidate the role of IL-17A in retinal capillary degeneration. Further, ex vivo studies were performed in retinal endothelial cells to identify the IL-17A-dependent mechanism that induces cell death. RESULTS: It was determined that diabetes-induced retinal capillary degeneration was significantly lower in IL-17A-/- mice. Further, retinal endothelial cell death occurred through an IL-17A/IL-17Râ¯ââ¯Act1/FADD signaling cascade, which caused caspase-mediated apoptosis. CONCLUSION: These are the first findings that establish a pathologic role for IL-17A in retinal capillary degeneration. Further, a novel IL-17A-dependent apoptotic mechanism was discovered, which identifies potential therapeutic targets for the early onset of diabetic retinopathy.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/fisiología , Diabetes Mellitus Experimental/complicaciones , Retinopatía Diabética/fisiopatología , Proteína de Dominio de Muerte Asociada a Fas/fisiología , Interleucina-17/fisiología , Vasos Retinianos/fisiopatología , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Capilares/fisiopatología , Caspasas/metabolismo , Muerte Celular , Diabetes Mellitus Experimental/fisiopatología , Células Endoteliales/fisiología , Proteína de Dominio de Muerte Asociada a Fas/genética , Proteína de Dominio de Muerte Asociada a Fas/metabolismo , Técnicas de Silenciamiento del Gen , Humanos , Interleucina-17/deficiencia , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
Diabetic retinopathy is a prevailing diabetes complication, and one of the leading causes of blindness worldwide. IL-17A is a cytokine involved in the onset of diabetic complications. In the current study, we examined the role of IL-17A in the development of retinal inflammation and long-term vascular pathology in diabetic mice. We found IL-17A expressing T cells and neutrophils in the retinal vasculature. Further, the IL-17A receptor was expressed on Muller glia, retinal endothelial cells, and photoreceptors. Finally, diabetes-mediated retinal inflammation, oxidative stress, and vascular leakage were all significantly lower in IL-17A-/- mice. These are all clinically meaningful abnormalities that characterize the onset of diabetic retinopathy.